In my previous study (Tee et al., Cell. 2014 Feb 13;156(4):678-90), I reported the importance of signal-induced chromatin remodeling as a key epigenetic priming event that underlies the transcriptional competent state of inactive developmental promoters in embryonic stem cells (ESCs). Developmental genes are typically inactive in ESCs, yet exhibit evidence of transcription initiation, i.e are `poised'. However, unexpectedly, these genes are devoid of TFIIH that comprises the canonical transcription initiation complex, suggesting that an alternative mechanism is employed to drive transcription initiation. My preliminary findings now show that DNA negative supercoiling (notably supported by the presence of topoisomerase 2A, TOP2A) may be key to the process, abrogating the function of TFIIH in transcription initiation. Strikingly, amongst all the developmental genes, neuronal genes are most conspicuously occupied by TOP2A, suggesting that neuronal genes may adopt distinct DNA topological and epigenetic configurations that are critical for their expression. Interestingly, recent exome sequencing studies have identified de novo mutations in topoisomerases in individuals with Autism Spectrum Disorder, epilepsy and intellectual disabilities. Therefore, these studies, together with my own, highlight an important, yet underappreciated role of DNA topology in neuronal gene regulation. In this proposal, I will explore the role of DNA topology during neural differentiation of ESCs, investigating how DNA supercoiling and topoisomerases act to promote robust expression of neuronal genes. The findings will help set the stage for determining how deregulation of the process may lead to neurodevelopmental and psychiatric disorders. I will employ novel high-throughput methodologies to probe for the dynamic changes in DNA topology in vivo, including the distribution of alternative DNA structures (i.e non-B DNA structures) that are formed as a function of DNA supercoiling. Notably, non-B DNA structures are notorious for their ability to induce genomic instability, and underlie more than 20 hereditary neurological diseases. Therefore, understanding where and how these structures are formed, and their impact on chromatin environment, is a necessary first step towards devising strategies for therapeutic intervention. Successful completion of this proposal will open up new vistas in understanding gene regulation, and will provide novel molecular insights into neurological disease etiologies. I have assembled a strong mentorship team including experts in the fields of DNA topology and neurobiology, as well as a comprehensive training plan that will facilitate my transition to independence.